Journal: The Journal of Biological Chemistry

Article Title: Wingless-type Mammary Tumor Virus Integration Site Family, Member 5A (Wnt5a) Regulates Human Immunodeficiency Virus Type 1 (HIV-1) Envelope Glycoprotein 120 (gp120)-induced Expression of Pro-Inflammatory Cytokines via the Ca 2+ /Calmodulin-dependent Protein Kinase II (CaMKII) and c-Jun N-terminal Kinase (JNK) Signaling Pathways *

doi: 10.1074/jbc.M112.381046

Figure Lengend Snippet: Wnt5a is critical for gp120 to induce IL-1β, IL-6, and TNF-α in the SDH. A–C, time courses of IL-1β (A), IL-6 (B), and TNF-α (C) protein levels after i.t. gp120 (100 ng) administration. D–F, inhibition of Wnt5a by Box5 impaired gp120-induced increase of IL-1β (D), IL-6 (E), and TNF-α (F) protein in the SDH at 12 h after gp120 injection. G–I, sFRP3 attenuated gp120-induced increase of IL-1β (G), IL-6 (H), and TNF-α (I) protein levels in the SDH at 12 h after gp120 injection. Box5 (10 μg), sFRP3 (2 μg) or vehicle was intrathecally administered 30 min prior to 100 ng of gp120 or vehicle injection. In summary graphs, the levels of target proteins were normalized against β-actin and expressed as relative units to the control (0 h or vehicle). Data presented in graphs are means ± S.E. from at least 3 mice per group from three experiments. *, p < 0.05; **, p < 0.01 versus control (one way ANOVA).

Article Snippet: The primary antibodies used included: rabbit anti-Wnt5a (0.5 μg/ml; cat. 72583; abcam), rabbit anti-Wnt5a (1 μg/ml; cat. 2530; CST), goat anti-IL-1β (0.2 μg/ml; cat. AF-401-NA; R&D systems), rat-anti-IL-6 (0.5 μg/ml; cat. 554400; BD Biosciences), goat-anti-TNF-α (0.1 μg/ml; cat. AF-410-NA; R&D systems), rabbit anti-p-αCaMKII (Thr-286) (1:1000; cat. 3361; Cell Signaling Technology (CST)), rabbit anti-CaMKII (1:1000; cat. 3362; CST), rabbit anti-p-JNK (Thr-183/Tyr-185) (1:1000; cat. 9251; CST), rabbit anti-JNK (1:1000; cat. 9252; CST), rabbit anti-p-PKC (pan) (Thr-514) (1:1000; cat. 9379; CST), and rabbit anti-β-actin (1:1000; cat. 4967; CST) antibodies.

Techniques: Inhibition, Injection

Journal: The Journal of Biological Chemistry

Article Title: Wingless-type Mammary Tumor Virus Integration Site Family, Member 5A (Wnt5a) Regulates Human Immunodeficiency Virus Type 1 (HIV-1) Envelope Glycoprotein 120 (gp120)-induced Expression of Pro-Inflammatory Cytokines via the Ca 2+ /Calmodulin-dependent Protein Kinase II (CaMKII) and c-Jun N-terminal Kinase (JNK) Signaling Pathways *

doi: 10.1074/jbc.M112.381046

Figure Lengend Snippet: Exogenous Wnt5a evokes the expression of IL-1β, IL-6, and TNF-α in the SDH. The time courses of IL-1β (A), IL-6 (B), and TNF-α (C) protein dynamics after i.t. Wnt5a (20 ng). In summary graphs, the levels of target proteins were normalized against β-actin and presented as relative units to 0 h. Data are expressed as means ± S.E. from at least three independent experiments with at least three animals. *, p < 0.05; **, p < 0.01 versus 0 h (one way ANOVA).

Article Snippet: The primary antibodies used included: rabbit anti-Wnt5a (0.5 μg/ml; cat. 72583; abcam), rabbit anti-Wnt5a (1 μg/ml; cat. 2530; CST), goat anti-IL-1β (0.2 μg/ml; cat. AF-401-NA; R&D systems), rat-anti-IL-6 (0.5 μg/ml; cat. 554400; BD Biosciences), goat-anti-TNF-α (0.1 μg/ml; cat. AF-410-NA; R&D systems), rabbit anti-p-αCaMKII (Thr-286) (1:1000; cat. 3361; Cell Signaling Technology (CST)), rabbit anti-CaMKII (1:1000; cat. 3362; CST), rabbit anti-p-JNK (Thr-183/Tyr-185) (1:1000; cat. 9251; CST), rabbit anti-JNK (1:1000; cat. 9252; CST), rabbit anti-p-PKC (pan) (Thr-514) (1:1000; cat. 9379; CST), and rabbit anti-β-actin (1:1000; cat. 4967; CST) antibodies.

Techniques: Expressing

Journal: The Journal of Biological Chemistry

Article Title: Wingless-type Mammary Tumor Virus Integration Site Family, Member 5A (Wnt5a) Regulates Human Immunodeficiency Virus Type 1 (HIV-1) Envelope Glycoprotein 120 (gp120)-induced Expression of Pro-Inflammatory Cytokines via the Ca 2+ /Calmodulin-dependent Protein Kinase II (CaMKII) and c-Jun N-terminal Kinase (JNK) Signaling Pathways *

doi: 10.1074/jbc.M112.381046

Figure Lengend Snippet: Wnt5a/CaMKII and Wnt5a/JNK pathways differentially regulate the expression of IL-1β, IL-6, and TNF-α in the SDH. A and B, effects of KN-93 (KN) and SP600125 (SP) on i.t. Wnt5a-evoked pT286-αCaMKII (A) and pT183/Y185-JNK (B) in the SDH. C–E, effects of KN-93, SP600125, and stearoylcarnitine (ST) on Wnt5a-induced IL-1β (C), IL-6 (D), and TNF-α (E) in the SDH. F–H, effects of KN-93, SP600125, and stearoylcarnitine (ST) on gp120-induced IL-1β (F), IL-6 (G), and TNF-α (H) in the SDH. KN-93 (25 μg), SP600125 (10 μg), and ST (20 μg) were intrathecally administered 30 min prior to i.t. Wnt5a (20 ng) or gp120 (100 ng). At 12 h after Wnt5a, gp120 or vehicle injection, the SDH was collected. In summary graphs, the levels of target proteins were normalized with the β-actin loading control and expressed as relative units to the vehicle. Data in the graphs are presented as means ± S.E. *, p < 0.05; **, p < 0.01; ***, p < 0.001 (one way ANOVA).

Article Snippet: The primary antibodies used included: rabbit anti-Wnt5a (0.5 μg/ml; cat. 72583; abcam), rabbit anti-Wnt5a (1 μg/ml; cat. 2530; CST), goat anti-IL-1β (0.2 μg/ml; cat. AF-401-NA; R&D systems), rat-anti-IL-6 (0.5 μg/ml; cat. 554400; BD Biosciences), goat-anti-TNF-α (0.1 μg/ml; cat. AF-410-NA; R&D systems), rabbit anti-p-αCaMKII (Thr-286) (1:1000; cat. 3361; Cell Signaling Technology (CST)), rabbit anti-CaMKII (1:1000; cat. 3362; CST), rabbit anti-p-JNK (Thr-183/Tyr-185) (1:1000; cat. 9251; CST), rabbit anti-JNK (1:1000; cat. 9252; CST), rabbit anti-p-PKC (pan) (Thr-514) (1:1000; cat. 9379; CST), and rabbit anti-β-actin (1:1000; cat. 4967; CST) antibodies.

Techniques: Expressing, Injection

Journal: Journal of Inflammation (London, England)

Article Title: Comparative effects of the herbal constituent parthenolide (Feverfew) on lipopolysaccharide-induced inflammatory gene expression in murine spleen and liver

doi: 10.1186/1476-9255-2-6

Figure Lengend Snippet: IL-6 protein production in sera following parthenolide and LPS co-treatment. Female B6C3F1 mice were co-treated with parthenolide (5 mg/kg, i.p.) or 50 μl DMSO and LPS (1 mg/kg, i.p.) or 100 μl water. After 90 minutes, blood was collected and serum analyzed for IL-6 by ELISA. The letter (a) indicates a significant difference compared to vehicle and parthenolide controls; (b) indicates a significant difference compared to LPS. Data are mean ± SEM (n = 16, controls n = 4), and is a combination of 4 separate experiments.

Article Snippet: IL-6 analysis was performed using purified and biotin-conjugated rat anti-mouse IL-6 antibodies from PharMingen (San Diego, CA) as described previously [ ].

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Journal of Inflammation (London, England)

Article Title: Comparative effects of the herbal constituent parthenolide (Feverfew) on lipopolysaccharide-induced inflammatory gene expression in murine spleen and liver

doi: 10.1186/1476-9255-2-6

Figure Lengend Snippet: IL-6 mRNA expression levels in spleen and liver following parthenolide and LPS co-treatment. Female B6C3F1 mice were co-treated with parthenolide (5 mg/kg, i.p.) or 50 μl DMSO and LPS (1 mg/kg, i.p.) or 100 μl water. Spleen and liver were collected after 90 minutes and total RNA was extracted and subjected to real-time, one-step RT-PCR using TaqMan primers and probes. IL-6 mRNA levels were normalized using 18S rRNA and related to spleen control values. (a) indicates a significant difference compared to vehicle and parthenolide controls; (b) indicates a significant difference compared to LPS. Data are mean ± SEM (n = 16, controls n = 4), and is a combination of 4 separate experiments.

Article Snippet: IL-6 analysis was performed using purified and biotin-conjugated rat anti-mouse IL-6 antibodies from PharMingen (San Diego, CA) as described previously [ ].

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction

Journal: Journal of Neuroinflammation

Article Title: Murine astrocytes produce IL-24 and are susceptible to the immunosuppressive effects of this cytokine

doi: 10.1186/s12974-019-1444-1

Figure Lengend Snippet: IL-24 augments the expression of suppressive cytokine signaling components by murine astrocytes and limits inflammatory cytokine release by these cells. a Astrocytes were untreated or treated with recombinant IL-24 (10, 30, or 100 ng/mL) for 30 min and the presence of phosphorylated STAT3 and STAT1 was determined by immunoblot analysis. Expression of β-actin is shown as a loading control and these immunoblots are representative of two separate experiments. b Astrocytes were untreated or treated with recombinant IL-24 (10, 30, or 100 ng/mL) for 2 or 4 h, and SOCS3 mRNA expression was determined by semi-quantitative RT-PCR. Expression of the housekeeping gene product GAPDH is shown, and relative SOCS3 expression was determined by densitometric analysis and normalized to untreated cells. Data is expressed as the mean ± the SEM of three independent experiments, and an asterisk indicates a statistically significant difference from unchallenged cells at each time point ( p < 0.05). c Astrocytes were untreated or treated with recombinant IL-24 (10, 30, or 100 ng/mL) for 8 h prior to immunoblot analysis for SOCS3 protein expression. Expression of the housekeeping gene β-actin is shown and relative SOCS3 protein expression was determined by densitometric analysis normalized to untreated cells. Asterisks indicate statistically significant differences from unchallenged cells (p < 0.05). d Astrocytes were untreated or treated with IL-24 (0.5, 3, 10, 30, or 100 ng/mL) for 4 h prior to challenge with bacterial LPS (5 ng/mL) or vehicle control for 12 h, and IL-6 secretion was determined by specific capture ELISA. Asterisks indicate a statistically significant difference ( p < 0.05) from similarly challenged cells in the absence of IL-24 ( n = 3). e Primary astrocytes were untreated or treated with recombinant IL-24 (10, 30, or 100 ng/mL) for 4 h prior to being uninfected or infected with Nm for 48 h before cell viability analysis via MTS assay. Data is presented as the mean absorbance ± SEM for three experiments. 0.1% Triton X-100 was used as a positive control, and an asterisk indicates a statistically significant difference from unchallenged cells in the absence of IL-24 ( p < 0.05)

Article Snippet: Commercially available Duoset® ELISA kits were used to measure IL-10 and TNF-α secretion (R&D Systems), while murine IL-6 secretion was measured using a rat anti-mouse IL-6 capture antibody (Clone MP5-20F3) and a biotinylated rat anti-mouse IL-6 detection antibody (Clone MP5-C2311) (BD Biosciences).

Techniques: Expressing, Recombinant, Western Blot, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Infection, MTS Assay, Positive Control